Telomere patterning at the chromosome level in matched donor and reprogrammed cell populations: biology, method development and investigation
Telomeres are dynamic structures composed of a DNA repeat, (TTAGGG)n in mammals, and associated proteins. Located at the very ends of each chromosome, they serve to shelter the terminus of the DNA strand and permit cellular proliferation. Telomere length, or the number of repeats, shortens with age and is also tied to a number of physiological, environmental, genetic and epigenetic factors. Despite significant length variation between the telomeres of different chromosome arms, the majority of past studies have been limited to comparisons of averaged values. This thesis presents a novel method, Component Distribution Comparison (CDC), which allows the quantitative and comprehensive comparison of chromosome arm specific telomere length. It is used to make comparisons between 11 somatic cell nuclear transfer (SCNT) clones, 5 presumptive induced pluripotent stem cell (iPSC) lines, 9 donors and 3 additional bovine animals. In a subset of 4 female t(X;23) animals, X-inactivation status and the associated telomere lengths of each X chromosome were also investigated. Overall, the chromosome arm specific telomere length characteristics measured in reprogrammed cells were significantly more similar to their donors than unrelated samples; however, discrepancies from complete matches between shared genome comparisons were also noted. This appears independent from a general elongation in telomere length in reprogrammed samples. These results suggest the telomere length at each chromosome arm is predominantly dictated by the genome, and is also impacted by other factors such as epigenetics or cellular history. An epigenetic and subtelomere interaction model is described; ‘outside-in’ and ‘inside-out’ frameworks to respectively assist in understanding telomere length determination and influence within an organism are also proposed. Significantly longer telomeres were noted on the active X vs. the inactive X of the t(X;23) donor animal. This divergence increased in 2 of 3 of her clones and abnormalities in X-inactivation were also noted. Together, the results of these studies demonstrate chromosome specific telomere length and X-inactivation carry significant potential to impact SCNT and iPSC outcomes and should be screened during reprogramming procedures. Further, they highlight the utility of both the described CDC method and reprogramming procedures to provide insights into telomere biology.